Whole-genome evolutionary dynamics of human parainfluenza virus type 3 in Shanghai, China, 2016–2024

Human parainfluenza virus type 3 (HPIV-3) is a major viral cause of acute respiratory infection (ARI), particularly in children, the elderly, and immunocompromised individuals. However, the limited availability of whole-genome data has impeded progress in viral evolution, vaccines, antiviral treatments, and effective preventive strategies. To address this gap, we conducted a comprehensive molecular evolutionary analysis of HPIV-3 based on whole-genome sequences obtained from clinical samples in Shanghai between 2016 and 2024. A total of 50 complete genomes were generated through a novel one-step amplification protocol combined with next-generation sequencing. Phylogenetic analysis revealed that all strains belonged to genotype C3. Time-scaled phylogenetic analysis estimated the tMRCA of HPIV-3 to be around 1925.55, earlier than previous estimates based on partial HN sequences, and revealed a stable population size prior to 1990 followed by two phases of sustained demographic expansion. Recombination events were detected in the HN gene and multiple other regions of the HPIV-3 genome. Entropy and selection pressure analyses demonstrated that the L gene showed the highest sequence variability and was the primary target of positive selection, emphasizing its role in viral evolution and adaptation. This study provides a comprehensive whole-genome–based evolutionary analysis of HPIV-3 in China and underscores the value of full-genome sequencing for accurate lineage classification, molecular surveillance, and a deeper understanding of viral dynamics. Our findings highlight the importance of integrating genomic data into epidemiological monitoring to inform the development of vaccines and antiviral strategies.